Cucurbit[7]uril (CB[7]) forms very stable complexes with the α-axial 5,6-dimethylbenzimidazole (α-DMB) nucleotide base when dissociated from the Co(III) center in vitamin B12 (CNCbl, KCB[7] = (7.5 ± 0.5) × 104 dm3 mol−1) and coenzyme B12 (AdoCbl, KCB[7] = (3.02 ± 0.35) × 106 dm3 mol−1). The inclusion of α-DMB ligand facilitates its release from the metal center and its subsequent protonation, with significantly higher pKbase-off values of 3.77 and 6.61, than determined for the free CNCbl (0.11) and AdoCbl (2.67), respectively. Addition of CB[7] to the base-on form of CNCbl at pH 2 provides for a direct measurement of the rate constant for the dissociation of the α-DMB ligand from the Co center (k = (4.6 ± 0.2) × 10−2 s−1, ΔH‡ = 93 ± 2 kJ mol−1, ΔS‡ = +41 ± 6 J K−1 mol−1). The β-axial 5′-deoxyadenosyl ligand (β-Ado) on coenzyme B12 is bound more weakly by a second CB[7] host (KCB[7] = (1.1 ± 0.2) × 103 dm3 mol−1), however inclusion of the β-Ado ligand has no effect on the kinetics of its heterolytic photodissociation from the Co(III) center.